Biology subjects

Liedtke, W.

Publications and source records attributed to Liedtke, W..

9 recordsLinked to original sources

bioRxiv2026

Acute Increase of Excitatory Activity in Pyramidal Neurons of Rat Motor Cortex under Static Magnetic Field

ObjectivesTranscranial application of a static magnetic field (SMF) was reported to result in subsequent modulation of neural excitability in the human motor cortex, but the acute mechanisms underlying this effect are unknown. We explore the mechanisms of this phenomenon with patch-clamp recording in rat brain slices during SMF exposure. Materials and MethodsPatch-clamp recording from layer II/III pyramidal neurons in motor cortex of acutely prepared brain slices were conducted during exposure to 0.20-0.35 T SMF or sham. ResultsDuring SMF exposure we observed an increase in the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) as well as miniature excitatory post-synaptic currents (mEPSCs) recorded in the presence of TTX to suppress action potentials and thus also network effects. There was a significant acute increase in sEPSC frequency for SMF exposure duration of both 6 min and 10 min, but not for 10 min sham exposure. After SMF exposure, the sEPSC and mEPSC frequency returned to baseline. The frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) was unaffected by SMF. The amplitude of the postsynaptic currents decreased with time for all recordings regardless of the condition, presumably due to the expected gradual deterioration of the patch clamp seal. ConclusionsThe acute effect of SMF exposure on sEPSC and mEPSC frequency, but not amplitude, is consistent with the assumption of a presynaptic or synaptic site mediating the effect. Furthermore, the consistency of the effect between sEPSCs and mEPSCs suggests that the effect is not related to action potential propagation in the presynaptic axon. The effect of SMF on EPSCs and not IPSCs may be related to the larger length of excitatory axons compared to inhibitory axons, or to effects on extracellular ionic gradients within the slice.

bioengineering↗

bioRxiv2025

Chondrocyte-Specific Knockout of Piezo1 and Piezo2 Protects Against Post-Traumatic Osteoarthritis Structural Damage and Pain in Mice

AbstractO_ST_ABSBackgroundC_ST_ABSOsteoarthritis (OA) is a debilitating joint disease characterized by cartilage degeneration, synovial inflammation, and bone remodeling, with limited therapeutic options targeting the underlying pathophysiology. Mechanosensitive ion channels Piezo1 and Piezo2 play crucial roles in chondrocyte responses to mechanical stress, mediating mechanotransduction pathways that influence chondrocyte survival, matrix production, and inflammatory signaling, but their distinct contributions to OA pathogenesis remain unclear. MethodsUsing inducible, chondrocyte-specific Aggrecan-Cre (Acan) mice, we investigated Piezo1, Piezo2, and combined Piezo1/2 conditional knockouts (cKOs) using the destabilization of the medial meniscus (DMM) model of post-traumatic OA in male and female mice. Pain and behavioral assessments were conducted at four time points to evaluate OA progression, while cartilage damage, bone remodeling, and synovial inflammation were assessed at the final endpoint of 28 weeks. Statistical analyses included one-way and two-way ANOVA with Tukeys multiple comparisons test. ResultsPiezo1 cKO delayed pain onset but ultimately exacerbated cartilage degradation and synovitis, emphasizing its dual role in protective and pathogenic mechanotransduction. While the Piezo2 cKO reduced pain and preserved activity, it failed to protect cartilage. Notably, Piezo1/2 cKO provided the greatest protection against cartilage degeneration, synovitis, and pain. Micro-computed tomography analyses revealed that Piezo2 is critical for maintaining trabecular bone integrity, with a Piezo2 cKO leading to decreased bone volume, thickness, and density, independent of injury. Piezo2 cKO also reduced normal meniscal ossification that occurs with age in mice. In contrast, a Piezo1/2 cKO normalized most bone remodeling parameters observed in Piezo2 cKO mice but did not restore medial tibial plateau thickness, highlighting Piezo2s essential role in bone structure. ConclusionsThese findings demonstrate the overlapping and compensatory roles of Piezo1 and Piezo2 in OA pathogenesis. Dual inhibition of Piezo1 and Piezo2 may offer a novel, effective therapeutic strategy targeting both structural and symptomatic aspects of the disease.

physiology↗

bioRxiv2021

Functional coupling between TRPV4 channel and TMEM16F modulates human trophoblast fusion

TMEM16F, a Ca2+-activated phospholipid scramblase (CaPLSase), is critical for placental trophoblast syncytialization, HIV infection, and SARS-CoV2-mediated syncytialization. How TMEM16F is activated during cell fusion is unclear. Here, we used trophoblasts as a model for cell fusion and demonstrated that Ca2+ influx through Ca2+ permeable transient receptor potential vanilloid channel TRPV4 is critical for TMEM16F activation and subsequent human trophoblast fusion. GSK1016790A, a TRPV4 specific agonist, robustly activates TMEM16F in trophoblasts. Patch-clamp electrophysiology demonstrated that TRPV4 and TMEM16F are functionally coupled within Ca2+ microdomains in human trophoblasts. Pharmacological inhibition or gene silencing of TRPV4 hindered TMEM16F activation and subsequent trophoblast syncytialization. Our study uncovers the functional expression of TRPV4 and a physiological activation mechanism of TMEM16F in human trophoblasts, thus providing us with novel strategies to regulate CaPLSase activity as a critical checkpoint of physiologically- and disease-relevant cell fusion events.

physiology↗

bioRxiv2021

Skeletal dysplasia-causing TRPV4 mutations suppress the hypertrophic differentiation of human iPSC-derived chondrocytes

Mutations in the TRPV4 ion channel can lead to a range of skeletal dysplasias. However, the mechanisms by which TRPV4 mutations lead to distinct disease severity remain unknown. Here, we use CRISPR-Cas9-edited human induced pluripotent stem cells (hiPSCs) harboring either the mild V620I or lethal T89I mutations to elucidate the differential effects on channel function and chondrogenic differentiation. We found that hiPSC-derived chondrocytes with the V620I mutation exhibited increased basal currents through TRPV4. However, both mutations showed more rapid calcium signaling with a reduced overall magnitude in response to TRPV4 agonist GSK1016790A compared to wildtype. There were no differences in overall cartilaginous matrix production, but the V620I mutation resulted in reduced mechanical properties of cartilage matrix later in chondrogenesis. mRNA sequencing revealed that both mutations upregulated several anterior HOX genes and downregulated antioxidant genes CAT and GSTA1 throughout chondrogenesis. BMP4 treatment upregulated several essential hypertrophic genes in WT chondrocytes; however, this hypertrophic maturation response was inhibited in mutant chondrocytes. These results indicate that the TRPV4 mutations alter BMP signaling in chondrocytes and prevent proper chondrocyte hypertrophy, as a potential mechanism for dysfunctional skeletal development. Our findings provide potential therapeutic targets for developing treatments for TRPV4-mediated skeletal dysplasias.

developmental biology↗

bioRxiv2021

Epidermal TRPV4 ion channels regulate UVB induced sunburn by triggering inflammmasome activation and MAPK signaling

Skin inflammation is an evolutionary-honed protective mechanism that serves to clear noxious cues and irritants and initiate regeneration. Calcium-permeable transient-receptor-potential (TRP) ion channels have critical functions in sensory transduction which is sensitized in skin inflammation. Skin sensory transduction relies on skin-innervating sensory neurons in the dorsal root ganglion (DRG), but also on innervated keratinocytes (KC). The multimodally-activated TRPV4 is robustly expressed in KC, where it can readily be activated by Ultraviolet-B (UVB). Our goal was to deconstruct keratinocyte TRPV4-mediated signaling, specifically how TRPV4 can facilitate inflammatory injury, thus lowering pain thresholds and rendering KC into pain-generator cells. We wanted to uncover the effect of TRPV4-mediated signaling on UVB-induced inflammasome activation in KC given the powerful impact of the activated inflammasome on pro-inflammatory/pro-algesic secretory signaling, using mouse models and cultured human KC. In mice, our evidence suggests that TRPV4 functions as calcium-permeable channel upstream of the KC inflammasome. Furthermore, we found that UVB induced activation of TRPV4 caused rapid - within minutes - Extracellular Signal Regulated Kinase (ERK) phosphorylation, caspase-1 activation and Interleukin-1{beta} (IL-1{beta}) secretion. In human primary KC we demonstrated that UVB induced secretion of IL-1{beta} was dependent on the NLR family pyrin domain containing 1 (NLRP1) inflammasome. Direct chemical TRPV4 activation could also activate NLRP1 and to lesser extent NLPR3. Building on our previous work, we now define at increased resolution TPRV4-dependent forefront signaling mechanisms in KC in response to UVB, showing TRPV4 upstream of the NLRP1 inflammasome, subsequent rapid ERK activation and pro-inflammatory/pro-algesic secretory function.

physiology↗

bioRxiv2020

Osteoarthritis: articular chondrocyte inflammatory signaling leads to enhanced gene expression and function of mechanotransduction channel Piezo1 as a pathogenic feedforwardmechanism

Osteoarthritis (OA) is a painful and debilitating condition of synovial joints without any disease-modifying therapies (1, 2). We previously identified mechanosensitive PIEZO channels, PIEZO1 and PIEZO2, both expressed in articular cartilage, to function in chondrocyte mechanotransduction in response to injury (3, 4). We therefore asked whether interleukin-1-mediated inflammatory signaling, as occurs in OA, influences Piezo gene expression and channel function, thus indicative of maladaptive reprogramming that can be rationally targeted. Primary porcine chondrocyte culture and human osteoarthritic cartilage tissue were studied. We found that interleukin-1 (IL-1) upregulated Piezo1 in porcine chondrocytes. Piezo1 expression was significantly increased in human osteoarthritic cartilage. Increased Piezo1 expression in chondrocytes resulted in a feed-forward pathomechanism whereby increased function of Piezo1 induced excess intracellular Ca2+, at baseline and in response to mechanical deformation. Elevated resting state Ca2+ in turn rarefied the F-actin cytoskeleton and amplified mechanically-induced deformation-microtrauma. As intracellular substrates of this novel OA-related inflammatory pathomechanism, in porcine articular chondrocytes exposed to IL-1 we discovered that enhanced Piezo1 expression depended on p38 MAP-kinase and transcription factors HNF4 and ATF2/CREBP1. CREBP1 directly bound to the proximal PIEZO1 gene promoter. In ensemble, these signaling and genetic re-programming events represent a novel and detrimental Ca2+-driven feed-forward mechanism that can be rationally targeted to stem the progression of OA. Significance StatementOsteoarthritis affecting weight-bearing joints is a global health problem, causing loss of mobility and enormous healthcare costs. Disease-modifying approaches are lacking. Here, we report a new cellular mechanism of inflammatory signaling in chondrocytes, the cellular substrate of cartilage. We show how osteoarthritis-relevant levels of interleukin-1 reprogram articular chondrocytes so that they become more susceptible to mechanical trauma, which chondrocytes sense via Piezo1/2 mechanosensitive ion channels. We uncover that IL-1 enhances gene expression of Piezo1 in primary articular chondrocytes underlying Piezo1 gain-of-function. We elucidate the new signaling pathway, from membrane to nucleus, including transcription factors that enhance Piezo1-expression. We also define detrimental effects of gain-of-function of Piezo1, for mechanotransduction and at-rest, that suggest this new reprogramming mechanism to contribute to osteoarthritis pathogenesis.

molecular biology↗

bioRxiv2020

A synthetic mechanogenetic gene circuit for autonomous drug delivery in engineered tissues

Mechanobiologic signals regulate cellular responses under physiologic and pathologic conditions. Using synthetic biology and tissue engineering, we developed a mechanically-responsive bioartificial tissue that responds to mechanical loading to produce a pre-programmed therapeutic biologic drug. By deconstructing the signaling networks induced by activation of the mechanically-sensitive ion channel transient receptor potential vanilloid 4 (TRPV4), we created synthetic TRPV4-responsive genetic circuits in chondrocytes. We engineered these cells into living tissues that respond to mechanical loading by producing the anti-inflammatory biologic drug, interleukin-1 receptor antagonist. Chondrocyte TRPV4 is activated by osmotic loading and not direct cellular deformation, suggesting tissue loading is transduced into an osmotic signal that activates TRPV4. Either osmotic or mechanical loading of tissues transduced with TRPV4-responsive circuits protected constructs from inflammatory degradation by interleukin-1. This synthetic mechanobiology approach was used to develop a mechanogenetic system to enable long-term, autonomously regulated drug delivery driven by physiologically-relevant loading.

synthetic biology↗

bioRxiv2019

Repurposing cell growth-regulating compounds identifies kenpaullone which ameliorates pathologic pain via normalization of inhibitory neurotransmission

Inhibitory GABA-ergic neurotransmission is fundamental for the adult vertebrate central nervous system and requires low chloride ion concentration in neurons. This basic ionic-homeostatic mechanism relies on expression and function of KCC2, a neuroprotective ionic transporter that extrudes neuronal chloride. Importantly, no other transporter can rescue KCC2 deficit, and attenuated expression of KCC2 is strongly associated with circuit malfunction in chronic pain, epilepsy, neuro-degeneration, neuro-trauma, and other neuro-psychiatric illnesses. To isolate Kcc2 gene expression-enhancing compounds, we screened 1057 cell growth-regulating compounds in cultured primary cortical neurons. We identified kenpaullone (KP), which enhanced Kcc2/KCC2 expression and function in cultured rodent and human neurons by inhibiting GSK3{beta}. KP effectively reduced pathologic pain in preclinical mouse models of nerve constriction injury and bone cancer. In nerve-injury pain, KP restored Kcc2 expression and GABA-evoked chloride reversal potential in the spinal cord dorsal horn. Delta-catenin, a phosphorylation-target of GSK3{beta} in neurons, activated the Kcc2 promoter via Kaiso transcription factor. Validating this new pathway in-vivo, transient spinal over-expression of delta-catenin mimicked KP analgesia. With relevance for pathologic pain, our discoveries of a newly repurposed compound and a novel genetically-encoded mechanism that each enhance Kcc2 gene expression enable us to re-normalize disrupted inhibitory neurotransmission through genetic re-programming.

neuroscience↗

bioRxiv2019

TRPV4 inhibition prevents increased water diffusion and blood-retina barrier breakdown in the retina of streptozotocin-induced diabetic mice.

A better understanding of the molecular and cellular mechanisms involved in retinal hydro-ionic homeostasis imbalance during diabetic macular edema (DME) is needed to gain insights into retinal physio(patho)logy that will help elaborating innovative therapies with lower health care costs. Transient receptor potential cation channel subfamily vanilloid member 4 (TRPV4) plays an intricate role in homeostatic processes that needs to be deciphered in normal and diabetic retina. Based on previous findings showing that TRPV4 antagonists resolve blood-retina barrier (BRB) breakdown in diabetic rats, we evaluated whether TRPV4 channel inhibition prevents and reverts retinal edema in streptozotocin(STZ)-induced diabetic mice. We assessed retinal edema using common metrics, including retinal morphology/thickness (histology) and BRB integrity (albumin-associated tracer), and also by quantifying water mobility through apparent diffusion coefficient (ADC) measures. ADC was measured by diffusion-weighted magnetic resonance imaging (DW-MRI), acquired ex vivo at 4 weeks after STZ injection in diabetes and control groups. DWI images were also used to assess retinal thickness. TRPV4 was genetically ablated or pharmacologically inhibited as follows: left eyes were used as vehicle control and right eyes were intravitreally injected with TRPV4-selective antagonist GSK2193874, 24 h before the end of the 4 weeks of diabetes. Histological data show that retinal thickness was similar in nondiabetic and diabetic wt groups but increased in diabetic Trpv4-/- mice. In contrast, DWI shows retinal thinning in diabetic wt mice that was absent in diabetic Trpv4-/- mice. Disorganized outer nuclear layer was observed in diabetic wt but not in diabetic Trpv4-/- retinas. We further demonstrate increased water diffusion and BRB hyperpermeability in diabetic wt mice, effects that were absent in diabetic Trpv4-/- mice. Retinas of diabetic mice treated with PBS showed increased water diffusion that was not inhibited by GSK2193874. ADC maps in nondiabetic Trpv4-/- mouse retinas showed restricted diffusion. Our data provide evidence that water diffusion is increased in diabetic mouse retinas and that TRPV4 function contributes to retinal hydro-ionic homeostasis and structure under control conditions, and to the development of BRB breakdown and increased water diffusion in the retina under diabetes conditions. A single intravitreous injection of TRPV4 antagonist is however not sufficient to revert these alterations in diabetic mouse retinas.

physiology↗